27. Deutscher Krebskongress

Artikel

The oncogenic potential of the homeobox gene Cdx2 is depending on the N-terminal transactivation domain in vivo and can be antagonized by inhibition of the MAPK pathway in a mouse model of t(12;13)(p13;q12) positive AML

Gliederung

We have recently shown that in a murine model of t(12;13)(p13;q12) AML the ectopic expression of the homeobox gene CDX2, induced by the chromosomal translocation, is the key event in myeloid leukemogenesis and not the expression of the ETV6-CDX2 fusion gene generated by the translocation (PNAS; 101; 2004). In an effort to characterize the functional relevance of different Cdx2 motifs for the transforming capacity of the gene we generated constructs, inactivating the DNA binding homeodomain (N51S−Cdx2), the PBX1−interacting motif (W167A−Cdx2), or deleting the N−terminal portion of Cdx2 (N−Cdx2). Expression of Cdx2 and the different mutants were retrovirally induced in primary murine bone marrow cells. The W167A−Cdx2 did not show a reduced activity in vitro compared to Cdx2 with an increase of primary colony formation (3−fold) (n=3;p<0.001) and a higher number of CFU−G/GM colonies (p<0.015) compared to the GFP control. In addition W167A−Cdx2 enhanced the replating capacity of clonogenic progenitors (80−100fold increase in secondary colonies (p<0.005)) and induced the outgrowth of blast colonies (2700fold; p<0.02). In contrast, the N51S−Cdx2 and the N−Cdx2 constructs lost the hematopoietic activity in vitro. In vivo all mice transplanted with cells expressing Cdx2 or the W167A−Cdx2 mutant developed transplantable AML. Of note, the N−Cdx2 mutant does not induced any leukemia in vivo (n=13). We extended structure-function analyses, inactivating the phosphorylation site (S60) in the Cdx2 transactivation domain, previously shown to be regulated by the MAPK family. The S60 mutant did not reduce the activity of Cdx2, but incubation with the MEK1 inhibitor PD98059 decreased the frequency of CFU-S 8fold (n=7; p<0.001) and blocked growth of leukemic Cdx2 transfected blasts in vitro, indicating that other phosphorylation sites are relevant for leukemic activity. These data demonstrate that the transforming activity of Cdx2 is depending on the N-terminal transactivation domain. Furthermore, our data link the oncogenic potential of Cdx2 directly to the MAPK signaling, opening the possibility to counteract Cdx2 associated leukemogenesis by kinase inhibitors.